Objective To observe dynamically the induction of reactive oxygen species (ROS) by titanium dioxide nanoparticles in the human epithelial cell line A549, which originated from a human lung carcinoma and composed of polarized alveolar type Ⅱ cells.
Methods The A549 cells were exposed to 10, 20, 40 μg/mL 5 nm diameter anatase TiO2 particles and <10nm(diameter)& #215;40nm(length)rutile TiO2 particles. Flow cytometry was used to analyze the fluorescence probes for ROS in the cells, consisting of Diehlo (DCFH-DA)located predominantly in cytosol, and Dihydrorhodamine 123 (DHR123)as mitochondriaassociated probe.
Results The biological response in cytoplasm and mitochondria to release ROS induced by the two types of TiO2 particles was similar. ROS could be detected as rapid as in 5 minutes after the exposure and could sustain for 120 min under tested concentrations (5-160μg/mL)and with dose dependent response. The peak of ROS was generated in 30 rain after 5 nm diameter anatase TiO2 particles treatment and 60 rain after <10nm(diameter)& #215;40 nm(length)rutile TiO2 particles treatment. Flow cytometric analysis showed that nanoparticles unexpectedly appeared in determination of ROS each time, and the stronger ROS fluorescence intensity the higher peak of nanoparticles could be observed. This result suggested that it might be possible to use a simple and easy method of flow cytometry to evaluate uptake potential of nanoparticles in mammalian cells and to replace transmission electron microscope (TEM) for quantifying nanoparticles in biological specimens.
Conclusion 5 nm diameter anatase and <10 nm (diameter)& #215;40 nm(length) ruffle TiO2 particles can induce ROS of cytosol and mitochondria in A549 cells.
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